The overall goal of this project is to learn about the interaction of domains in non-ribosomal polypeptide synthetase modules, using the 142 kDa E. coli EntF. This protein consists of four domains, C, A, T and TE that act as chain elongation and termination module to make and release the enterobactin siderophore. We are interested in how the domains cooperate to synthesize enterobactin, and how the HS-pantetheinyl-P group bound to the TE domain affects the interaction of domains, and how this may relate to the assembly- line function of the protein. The multidomain protein is a model system for development of NMR for large proteins The specific aims are: Specific Aim 1.. Complete structure determination of the 36 kDa A-T didomain fragment of the 142kDa EntF Specific Aim 2 Studies of structures and interactions of the in cis C and A domains centering on the T domain scaffold and how these partner proteins dock with it SpecificAim 3. Studies of the structure of the A-T 70 kDa didomain of EntF Specific Aim 4. . Interaction of the HS-pantetheinyl-P prosthetic group (isotopically labeled or spin-labeled )on the two T domains in EntB and EntF with different domains in cis and in trans (C, A, TE) in unacylated and acylated forms